Invertebrates have experienced intense selection pressure for sensitive and effective ability to locate mates, food and hosts in nature. For example, Microplitis croceipes Cresson (Hymenoptera: Braconidae) are solitary insect endoparasitoids of the larvae of Heliocoverpa zea Boddie and Heliothis virescens Boddie and are able to learn and respond to volatile chemicals in amounts in at least the pico gram/s range (Wäckers et al., Journal of Food Science, Volume 76, 41-47, 2011). Such low detection thresholds are comparable to those of vertebrates (Smith et al., Annu. Rev. Entomol., Volume 39, 351-375, 1995; Stoddart, In: The Ecology of Vertebrate Olfaction, 58-62, 1980, Chapman and Hall, New York, N.Y.). The fact that invertebrates are able to learn allows them to be programmable. The breadth of invertebrate learning abilities has increasingly been studied and described (Fedodov, V P, J. of Evolutionary Biochemistry and Physiology, volume 45, 1-26, 2000; Willows et al., In. Invertebrate Learning. Plenum Press, New York, 1973-1975; Abramsom, C I, In. Invertebrate Learning, Washington D.C., American Psychological Assn., 1990; Zang, W., Environmental Monitoring and Assessment, volume 130, 415-422, 2007; Papaj et al., In. Insect Learning. Ecological and Evolutionary Perspectives. Chapman and Hall, 1993; Vet et al., In: Chemical Ecology of Insects 2, 65-101, 1995, Chapman and Hall, New York, N.Y.; Menzel et al., In. Biology of Learning: Report of the Dahlem Workshop on the Biology of Learning, 249-270, Springer-Verlag, Berlin, 1983).
One approach in detecting compounds is to utilize conditioned invertebrates to detect the presence of a compound. One such model invertebrate is the insect species, Microplitis croceipes, as disclosed in U.S. Pat. No. 6,919,202 and U.S. Pat. No. 7,607,338 and incorporated herein by reference. U.S. Pat. No. 6,919,202 discloses a method of training an organism to detect at least one chemical by repeated exposure of a biological resource in the presence of a target chemical. U.S. Pat. No. 7,607,338 discloses a similar method to detecting the presence of at least one chemical wherein a trained insect is utilized in conjunction with a portable handheld apparatus. However U.S. Pat. Nos. 6,919,202 and 7,607,338 are limited in that the objective of the training is the detection of the conditioned chemical compound or blend of compounds, rather than the detection of a particular concentration. For the actual detection of particular concentrations, or ranges of concentrations (above or below a threshold), there is a need to further investigate conditioning methods in which the organism to be conditioned is conditioned at the threshold concentration or at a range of concentrations of a particular compound.
There is a crucial need to develop an invertebrate biosensor based on an organisms' ability to detect and report specific concentrations of targeted compounds. The detection of a compound of interest at and beyond a threshold concentration is particularly relevant in a number of applications, for example the monitoring of food quality, plant-, animal- and human-health. Although there are few studies of odor concentration learning in invertebrates, several studies of honeybees, Apis mellifera (Hymenoptera: Apidae) (Marfaing et al., J. Insect Physiol., volume 35, 949-955, 1989; Bhagavan and Smith, Physiol. Behav., volume 61, 107-117, 1996; Wright et al., Proc. R. Soc. B 271: 147-152, 2004; but see Pelz et al., J. Exp. Biol. 200: 837-847, 1997) found that learning and detection of odors is concentration dependent. Marfaing et al. (J. Insect Physiol., volume 35, 949-955, 1989) and Wright et al. (Proc. Royal Soc. B. volume 272, 2417-2422, 2005) also showed that concentration learning and detection of odors in the honeybee is compound dependent, and Wright et al. (J. Comp. Physiol. A 191: 105-114, 2005) further showed that learning and detection of compounds in mixtures depends on the concentration and the ratio (proportion) of the compounds in a mixture. Kaiser and De Jong (Animal Learning & Behavior 23: 17-21, 1995), found that the parasitoid Leptopilina boulardi (Hymenoptera: Figitidae) responds to absolute and relative concentrations of banana and strawberry odors in accordance with their conditioning concentration.
In one particular application, use of a conditioned invertebrate can detect a broad range of concentrations of a compound related to boar taint. Boar taint is an offensive odor or taste emanating from pork products. The boar taint is derived from non-castrated male pigs and the major chemicals responsible for boar taint are skatole and androstenone (Lundstrom K, et al., 2009, Animal 3:1497-1507). Skatole (3-methyl-indole) and androstenone (5α-androst-16-en-3-one) are fat soluble and accumulated in fat tissue of male pigs upon reaching sexual maturity. As such, if the concentration of skatole in adipose tissue of entire male pig exceeds 0.2 μg/g to 0.25 μg/g, consumers can perceive the tainted pork product. (Id.) For androstenone, consumers can perceive tainted pork products when androstenone is between 0.5 μg/g to 1.0 μg/g for the entire carcass. (Id.)
Methods to control for boar taint include castrating male pigs at an early age before reaching sexual maturity, selective breeding of only female pigs, selective breeding of pigs that are known to have low boar taint, or slaughtering pigs at an early age (approximately 6 months in age) so the skatole and androstenone do not accumulate in pre-adolescent adipose tissue.
However, the method of castrating male pigs have brought questions of animal welfare concerns. Specifically, in the European Union animal welfare concerns during the castration process has moved the EU to seek alternatives to castration. Furthermore, the EU has set a goal to end surgical castration of pigs in the future. Given that boar castration may be limited, there is a need to develop a low cost and rapid post-slaughtering methodology to detected boar taint.
There is a need to further investigate invertebrate ability to learn odor concentrations and whether the invertebrate response is compound dependent and incorporating those compound dependent responses to detect a range of compounds.